On the paradoxically concentration-dependent metabolism of 6-mercaptopurine in WEHI-3b murine leukemia cells.
The intracellular metabolism of 6-mercaptopurine (6-MP) was studied in a murine leukemia cell line, WEHI-3b. Cells were incubated 3 to 24 h with 10 nM to 50 microM 6-MP. Nucleotides were extracted with perchloric acid, and the 6-thiopurine nucleotides were isolated on mercurial cellulose. The endogenous ribonucleotides in the perchloric acid extracts as well as 6-thiopurine nucleotides were separated and quantified with anion exchange high-performance liquid chromatography. The concentration of 6-thioinosinate (6-TIMP) and 6-thioxantinate (6-TXMP) increased with an increasing 6-MP dose. The concentration of the 6-thioguanosine nucleotides (6-TGN) increased with 6-MP concentrations between 10 nM and 1 microM. However, further increase in 6-MP concentration led to a decrease in the formation of 6-TGN. At 50 microM 6-MP, the concentration of 6-thioguanosine 5'-triphosphate was one fifth of that seen at 1 microM. The incorporation of 6-[35S]mercaptopurine into DNA was also slightly higher at 1 microM compared with 50 microM. The cytocidal effect on clonogenic cells was one log greater at 1 microM 6-MP compared with 50 microM 6-MP. The decrease of 6-TGN was accompanied not only by an increased 6-TIMP concentration but also by an inhibition of the purine de novo synthesis and consequently by a decrease of the cellular ATP concentration. The ATP concentration in the cells treated with 1 microM 6-MP could be reduced to the level seen in cells treated with 50 microM 6-MP by simultaneous incubation with 0.3 microM antimycin A. This decrease of ATP concentration was accompanied by a reduction of 6-TGN and to a lesser extent of 6-TXMP. These experiments suggest that the "self-limiting" phenomenon in the metabolism of 6-MP might be caused by a depletion of ATP by inhibition of purine de novo synthesis presumably by 6-TIMP.
Liliemark
J
J
Department of Medicine, Karolinska Institute at Huddinge Hospital, Sweden.
Pettersson
B
B
Engberg
B
B
Lafolie
P
P
Masquelier
M
M
Peterson
C
C
D000818:Animals; D001711:Biotransformation; D002250:Carbon Radioisotopes; D002460:Cell Line; D007700:Kinetics; D007942:Leukemia, Experimental / Q000378:metabolism*; D015122:Mercaptopurine / Q000378:metabolism* / Q000494:pharmacology; D051379:Mice; D014407:Tumor Cells, Cultured / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D014410:Tumor Stem Cell Assay
D002250:Carbon Radioisotopes; D015122:Mercaptopurine